title: Performance evaluation of affinity ligands for depletion of abundant plasma proteins.
abstract: Human plasma is a commonly used diagnostic fluid in clinical chemistry. In-depth plasma proteomic analysis is performed to search for disease biomarkers, however the large dynamic range of protein abundance in plasma presents a substantial analytical challenge. Removal of abundant plasma proteins using antibody capture approaches is a common and attractive means to reduce sample complexity and to aid the analysis of lower abundance proteins of interest. A novel class of heavy chain camelid-derived affinity ligands produced in Saccharomyces cerevisiae, has recently been developed as an alternative to antibody-based depletion methods. Here, we evaluate the performance characteristics of these ligands for removal of high abundance plasma proteins. Affinity ligands were tested for the removal of 14 abundant human plasma proteins. The performance characteristics were evaluated by gel-electrophoresis and LC-MS/MS of the bound and flow-through fractions. The capacity of a 5.6mL column was found to be 125μL of plasma. Replicate analysis demonstrated high column reproducibility and linearity, efficient removal of abundant proteins, and enrichment of lower abundance proteins observed after depletion. The novel class of affinity ligands provides an attractive alternative to traditional antibody-based immunodepletion methods.
authors.affiliation: Canary Center at Stanford for Cancer Early Detection, Department of Radiology, Stanford University School of Medicine, Palo Alto, CA, USA.
authors.forename: Majlinda
authors.initials: M
authors.lastname: Kullolli
authors.forename: Jonathan
authors.initials: J
authors.lastname: Warren
authors.forename: Maria
authors.initials: M
authors.lastname: Arampatzidou
authors.forename: Sharon J
authors.initials: SJ
authors.lastname: Pitteri
mesh_terms: D001798:Blood Proteins / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D002846:Chromatography, Affinity / Q000379:methods*; D006801:Humans; D008024:Ligands; D020543:Proteome / Q000032:analysis; D040901:Proteomics / Q000379:methods*; D011994:Recombinant Proteins / Q000378:metabolism; D015203:Reproducibility of Results; D012441:Saccharomyces cerevisiae; D053719:Tandem Mass Spectrometry
chemical_list: D001798:Blood Proteins; D008024:Ligands; D020543:Proteome; D011994:Recombinant Proteins
keywords: Affinity ligands; Immunoaffinity/immunodepletion alternative; Ligand affinity; Plasma proteomics; Protein depletion